In 1813, Vautier reported the apparent cure of cancer in patients who concurrently suffered from gas gangrene caused by the infection of the anaerobic bacterium Clostridium perfringens. Since then, extensive studies have been undertaken on the deliberate injection of various bacteria including Salmonella typhimurium in addition to Clostridium, with the concept of bacteria-mediated cancer therapy. As a cancer grows over a certain size (approximately 1 mm3), it undergoes an undersupply of oxygen from surrounding vessels, with the formation of hypoxia within the cancer tissue. The cancer tissue in a hypoxia state provides an environment in which anaerobic bacteria are readily likely to proliferate while they might attack surrounding cancer cells to necrosis using various toxins and/or through unknown mechanisms. Various attempts have been made to enhance bacteria-mediated cancer therapy. For example, bacteria are transformed to express anticancer proteins, or are used to deliver a plasmid carrying an anticancer protein gene to a cancer tissue. To mitigate the side effects arising from bacterial proliferation and toxins, auxotrophic mutants, spores, and attenuated bacteria have also been tried for cancer therapy. In spite of these efforts, the use of bacteria in cancer therapy always has the risk of bacterial proliferation in normal tissues or organs surrounding the cancer mass.
There are broadly speaking two different types of cell wall in bacteria, called Gram-positive and Gram-negative. Gram-negative bacteria, such as Escherichia coli, Neisseria meningitidis, Pseudomonas aeruginosa, and Shigella flexneri, are known to spontaneously shed microvesicles from the outer membrane. The Gram-negative bacterial cell-derived shedding microvesicles are known as outer membrane vesicles typically consisting of a lipid bilayer. They are generally spherical with a size of 20˜200 nm, and have various biologically active substances, such as lipopolysaccharide (LPS), and outer membrane proteins, lipids and genetic materials (DNA, RNA) which influence the inflammatory responses of host cells. Bacterial cell-derived shedding microvesicles serve as an information carrier which plays a role in the transport of proteins or genetic materials between homogeneous cells and in cell-to-cell signaling, and contributes to the removal of competitive organisms or the survival of bacteria. In addition, the shedding microvesicles deliver toxins to hosts, thus accounting, in part, for the etiology of bacterial diseases. Reports that shedding microvesicles were found in the blood of patients who died of severe sepsis suggest that shedding microvesicles play an important role in the pathology of sepsis, which is characterized by systemic inflammation. This is also supported by the research finding that bacterial cell-derived shedding microvesicles stimulate host cells to secrete inflammatory cytokines and coagulants.
Gram-positive bacteria including Bacillus subtilis and Staphylococcus aureus produce shedding microvesicles, which was first found by the present inventors. However, there is a need for more information on components or functions of Gram-positive bacterial cell-derived shedding microvesicles.
Microvesicles that spontaneously shed from various bacteria species have been isolated and observed. Typically, shedding microvesicles are isolated from cell cultures by filtration or ultracentrifugation. In addition, it is known that the production of shedding microvesicles can be controlled with an antibiotic such as gentamicin. Particularly, there is a suggestion that shedding microvesicles prepared by treatment with a detergent might be applied as a vaccine against the infection of N. meningitidis, a bacterial pathogen. However, the capacity of these production methods is seriously limited.
In spite of the relationship between cancer and bacteria, there have been no reports on the role of bacterial cell-derived shedding microvesicles in the onset and progression of cancer, particularly, on the application of bacterial cell-derived shedding microvesicles to the treatment and/or diagnosis of cancer, thus far.